There is a fundamental gap in understanding how diabetes impacts the retinal neurons that provide vision. Because of this, current therapies intervene in latter stages of the disease, when blood vessels are affected. Anti-VEGF therapy limits damage in advanced diabetic retinopathy (DR), but does not address damage to the neurosensory retina. Hence, the long-term goal of this project is to define the mechanisms that cause retinal ganglion cell (RGC) dysfunction in patients with DR. The overall objective is to define the roles of mechanistic target of rapamycin complex 2 (mTORC2) signaling in the interactions between hyperglycemia and protein synthesis that lead to RGC dysfunction and subsequent vision loss in DR. The project will utilize diabetic rats and mice, which are the standard pre-clinical models of DR, and which exhibit the early neurodegenerative changes of human DR. The central hypothesis is that diabetes-induced defects in mTORC2 signaling impair protein synthesis, axonal function and survival of retinal ganglion cells. The rationale for this work is that identifying pathways causing RGC dysfunction and death will ultimately lead to preventive treatments and better vision for persons with diabetes. The hypothesis is supported by strong preliminary data showing: 1) prominent mTOR and Rictor expression in RGCs; 2) reduced mTOR expression in diabetic human donor eyes; and 3) the ability to conditionally knockout of Rictor expression in adult mice. The hypothesis will be tested in two specific aims: 1) to define the mechanisms by which diabetes impairs retinal mTORC2 activity and the cell-specific effects of diabetes on protein synthesis, and 2) to define the consequences of impaired mTORC2 activity on retinal ganglion cell protein synthesis, survival and morphology. The first aim will examine the effect of diabetes on mTORC2 complex member protein expression in human retinas, use diabetic rodent models to determine the molecular mechanisms by which diabetes reduces retinal mTORC2 activity and protein synthesis, and examine the cell-specificity of these effects. The second aim will employ in vivo spatial and cell- specific targete loss-of-function studies to disrupt mTORC complexes by knockout of the mTOR-associated proteins Rictor and Raptor in inner retinal neurons and determine the effects on protein turnover, axonal function and cell survival. The proposal is innovative because it: 1) expands the novel observation of diminished mTORC2 activity in DR and investigates the unexplored area of mTOR function in the neural retina; 2) will be the first to examine the cell-specific alterations i retinal mTORC complexes and protein synthesis leading to loss of neuronal integrity in DR; and 3) will use innovative techniques, including translatomics to define cell-specific changes in retinal mRNA translation, and targeted AAV-cre recombinase vectors to delete Rictor, Raptor and protein phosphatase PP2A genes in RGC. The work is significant because it will elucidate clinically relevant means to restore defective signaling pathways responsible for loss of RGC function in retinal diseases that have universal importance to vision and implications for the NEI Audacious Goals Initiatives.